A Novel Culturing Chip (cChip) Can Facilitate Culturing of Unculturable Bacteria From Aquatic Environment

Abstract Background: Culturing the unculturable microorganisms is an important aspect of microbiology. Once cultured the unculturable microorganisms can be a source of useful antibiotics, enzymes etc. Several studies have been conducted on culturing the unculturable microorganisms from soil. But little knowledge exists about culturing such bacteria from aquatic environment. Therefore, in this study we designed a novel culturing chip (cChip) to facilitate the growth of unculturable aquatic bacterial community. cChip was optimized for microbial growth using known bacteria in the lab, later, microbes from a freshwater lake were concentrated (instead of using the traditional dilution method) and inoculated in cChip before incubating the chip in simulated lake environment. Then further sub-culturing was done on laboratory media. The field samples were also analyzed using traditional culturing and metagenomics for a comparison. Results: Metagenomics analysis showed that 832 microbial species were present in the samples belonging to five different phyla, that is, Cyanobacteria, Proteobacteria, Actinobacteria, Bacteroidetes and Verrucomicrobia. However, traditional culturing yielded only 36 isolates that belonged to four different phyla, that is, Actinobacteria, Proteobacteria, Firmicutes and Bacteroidetes. Culturing through cChip yielded 154 isolates belonging to five different phyla, that is, Proteobacteria, Actinobacteria, Bacteroidetes, Verrucomicrobia and Firmicutes. Out of these 154 cChip isolates, 45 were previously uncultured bacteria having a 16S rRNA gene similarity from 91.35 % to 98.7 % to their closest relatives according to NCBI GenBank. Conclusion: This study shows that culturing microorganisms in the cChip from aquatic environment using concentration process before performing the traditional petri plate culturing can result in the successful growth of unculturable bacteria. To the best of our knowledge this is the first study conducted for successful exploration of unculturable aquatic microbial community. This study can have a significant impact on our understanding of the techniques that can be applied for exploring the unexplored microbiome from diverse environments. We also hypothesize that certain modifications in the manufacturing material, keeping the design and techniques of this study intact can result in the application of this technique from gut microbiome to extremophiles in the environment.